Retroviruses are the causative agents of a large variety of diseases including leukemias and lymphomas, solid tumors and immunosuppressive disorders in animals and humans. Understanding the details of viral replication and viral assembly is essential for development of new types of anti-retroviral therapies. In addition, potentially pathogenic retroviruses have been developed as useful viral vectors, both for research and for possibly for human gene therapies. Foamy viruses (FV) are a group of nonpathogenic retroviruses which have not been extensively studied. These viruses have potential as efficient and safe gene therapy vectors. The first specific aim of this proposal includes experiments aimed at understanding the details of the recognition signals in both RNA and protein that lead to specific genomic encapsidation. Over the past funding period we have shown that the alpharetroviruses such as RSV provide the most clearly defined system to understand retroviral genomic packaging. A small contiguous stretch of RNA serves as a sufficient signal for RNA encapsidation. We wish to continue this work with the hope of determining the tertiary structure of the RSV packaging RNA, and to understand the protein residues in nucleocapsid which are involved in genomic RNA recognition and viral assembly. The second and third specific aims are designed to understand how FV assemble into particles and package their enzymes. We will use a molecular genetic approach to understand the Gag-Gag, Gag-Pol and Gag-Pol-RNA interactions necessary for assembly. Foamy viruses bridge the replication pathways of retroviruses and the human pathogen hepatitis B virus. Studies of how the FV Gag and Pol genes are assembled into particles, and how the uniquely expressed Pol protein is regulated, will increase our knowledge of the possible ways retroviruses replicate, and provide interesting contrasts to both conventional retroviruses and hepatitis B virus.